Preparation And Tissue Structure Analysis Of Horse Bone Collagen Peptide

In this study,horse bones with different ages were selected as experimental materials.Bone tissue staining method was used to study the distribution of collagen fibers and calcium deposition in horse bone.The degreasing of horse bone was optimized by ultra-high pressure method.Acid-enzyme binding-method was used to extract collagen from horse bone,so as to prepare collagen peptides.The antioxidant activity of collagen peptides was analyzed.The results are as follows:The collagen content and calcium salt distribution characteristics in horse bone were observed by staining and image analysis.The results showed that the collagen fibers of 13-year-old horse horse bones were loose,disordered and uneven in density distribution,and there was significant difference between 2-year-old horse bones and 8-year-old horse bones(P < 0.05),but there was no significant difference between 2-year-old horse bones and 8-year-old horse bones(P > 0.05).Compared with other domestic animal bones such as cattle and sheep,the collagen fibers of adult horse bones were neat and regular in shape,uniform in density,orderly in arrangement,dense and high in bone density.Ultra-high pressure was used to assist the degreasing of horse bone powder with ethyl acetate solution,and the degreasing conditions were optimized by single factor and orthogonal test.The obtained technological conditions were as follows: ultra-high pressure300 MPa,holding time 9min,solid-liquid ratio 1:15g/m L.Under these conditions,the degreasing rate of horse bone reached 73.56%.Equine bone collagen was extracted by acidase binding and its structural characterization was studied.The thermal stability analysis of equine bone collagen of different ages showed that the denaturation temperature of equine bone collagen of different ages increased slightly with age.The thermal denaturation temperature of equine bone collagen after ultra-high pressure treatment is slightly lower than that of horse bone collagen treated in the traditional way.This indicates that the collagen crosslinking bond in the bone meal after ultra-high pressure treatment is broken,and the stability is weakened,resulting in a decrease in thermal stability.The effect of alkaline protease on the extraction rate of collagen peptides was studied.4 factors including enzymatic hydrolysis temperature,enzymatic hydrolysis time,p H value and enzyme dosage were screened.The optimum enzymatic hydrolysis conditions of horse bone collagen were obtained by orthogonal experiment as follows: enzymatic hydrolysis temperature for 55℃,p H=9.0,enzyme dosage for 4%,enzymatic hydrolysis time for 4h,and the yield of short peptides under this condition was 66.24%.Collagen peptides were separated by ultrafiltration device,and collagen peptide with molecular weight of 5-10 KDa,3-5KDa,1-3KDa and < 1KDa were obtained.In addition,their antioxidant effects were analyzed.The results showed that the antioxidant activity of collagen peptides is increased with increasing concentration.Among 4 peptides with different molecular weights obtained by ultrafiltration,the antioxidant activity of collagen peptide with molecular weight of 1-3KDa was significantly higher than that of other collagen peptides(P < 0.05).Under the condition of the same molecular weight,the antioxidant activity of collagen peptides for horse bones with different ages was different,and the antioxidant activity of 8-year-old horse bone was stronger.